Biodegradability of Disinfectants in Surface Waters from Buenos Aires: Isolation of an Indigenous Strain Able to Degrade and Detoxify Benzalkonium Chloride

  • María Susana Fortunato
  • Sabina Baroni
  • Ana Julieta González
  • Jesús David Álvarez Roncancio
  • Mariana Papalia
  • Manuela Martinefsky
  • Valeria Trípodi
  • Estela Planes
  • Alfredo Gallego
  • Sonia Edith Korol


Biodegradability of chlorhexidine (CH), triclosan (TC), and benzalkonium chloride (CBA) has been tested in 18 surface water sampling points in the urban area of Buenos Aires. Sampling points were located in both the Reconquista and the Matanza-Riachuelo basins as well as in the La Plata River. High tolerance to the three disinfectants was found and indigenous strains capable of degrading CBA and TC were isolated. Neither tolerance nor biodegradation were correlated with sewage pollution. A strain that degrades CBA was identified as belonging to the genus Pseudomonas using the API20NE system and 16SRNA sequencing. In batch assays, the strain was capable of degrading 100, 200, and up to 500 mg L−1 of CBA in 10, 25, and 46 h respectively with specific growth rates (μ) of 0.56, 0.30, and 0.14 h−1. The efficiency of the process was between 99.5–98.0% in terms of compound removal and between 93.8–89.1% in terms of chemical oxygen demand (COD). The detoxification of the compound as a result of the biodegradation was assessed using Pseudokirchneriella subcapitata, Vibrio fischeri, and Lactuca sativa as test organisms.


Benzalkonium chloride Triclosan Chlorhexidine Pseudomonas Biodegradation Detoxification 



We also thank Dr. Gabriel Gutkind and Dr. Marcela Radice from the Laboratory of Bacterial Resistance, Facultad de Farmacia y Bioquímica, Universidad de Buenos Aires, for his collaboration in the identification of microorganisms.

Funding Information

We thank the University of Buenos Aires for the grant given for this study, supported by UBACYT Program-Project 20020130100378BA 2014-2017.


  1. ACUMAR (2015). Base de datos hidrológica-Cuenca Matanza Riachuelo. BDH-CMR Accessed 27 March 2017.
  2. Al-Ahmad, A., Wiedmann-Al-Ahmad, M., Schön, G., Daschner, F. D., & Kümmerer, K. (2000). Role of Acinetobacter for biodegradability of quaternary ammonium compounds. Bulletin of Environmental Contamination and Toxicology, 64(6), 764–770. Scholar
  3. Alexander, M. (1999). Biodegradation and bioremediation (2nd ed.). London: Academic Press.Google Scholar
  4. APHA-American Public Health Association, American Water Works Association, Water Environment Federation. (2012). Standard methods for the examination of water and wastewater (22nd ed.). Washington, DC: APHA.Google Scholar
  5. Baroni, S., Calcagno, M. L., Fortunato, M. S., Álvarez Roncancio, J. D., Gallego, A, & Korol, S. E. (2016) Evaluación de un método colorimétrico directo para la medición de cloruro de benzalconio. The Journal of the Argentine Chemical Society 103 (1–2), Accessed 10 February 2017.
  6. Clara, M., Scharf, S., Scheffknecht, C., & Gans, O. (2007). Occurrence of selected surfactants in untreated and treated sewage. Water Research, 41(19), 4339–4348. Scholar
  7. Dann, A. B., & Hontela, A. (2011). Triclosan: environmental exposure, toxicity and mechanisms of action. Journal of Applied Toxicology, 31(4), 285–311. Scholar
  8. Das, R., Ghosh, S., & Bhattacharjee, C. (2015). A green practice for pharmaceutical drug chlorhexidine digluconate treatment and ecotoxicity assessment. Journal of Water Process Engineering, 7, 266–272. Scholar
  9. Ding, W. H., & Liao, Y. H. (2001). Determination of alkylbenzyldimethylammonium chlorides in river water and sewage effluent by solid-phase extraction and gas chromatography/ mass spectrometry. Analytical Chemistry, 73(1), 36–40. Scholar
  10. Dougherty, J. A., Swarzenski, P. W., Dinicola, R. S., & Reinhard, M. (2010). Occurrence of herbicides and pharmaceutical and personal care products in surface water and groundwater around Liberty Bay, Puget Sound, Washington. Journal of Environmental Quality, 39(4), 1173–1180. Scholar
  11. Dynes, J. J., Lawrence, J. R., Darren, R., Korber, D. R., Swerhone, G. D. W., Leppard, G. G., et al. (2006). Quantitative mapping of chlorhexidine in natural river biofilms. Science of the Total Environment, 369(1–3), 369–383. Scholar
  12. Ferrer, I., & Furlong, E. T. (2001). Identification of alkyl dimethylbenzylammonium surfactants in water samples by solid-phase extraction followed by ion trap LC/MS and LC/MS/MS. Environmental Science and Technology, 35(12), 2583–2588. Scholar
  13. Freitag, D., Geyer, H., Kraus, A., Viswanathan, R., Kotzias, D., Attar, A., et al. (1982). Ecotoxicological profile analysis: VII. Screening chemicals for their environmental behavior by comparative evaluation. Ecotoxicology and Environmental Safety, 6(1), 60–81 0147–6513/82/010060–22$02.00/0.CrossRefGoogle Scholar
  14. EPA (Environmental Protection Agency) (1989). Protocols for short term toxicity screening of hazardous waste sites. A.8.7. Lettuce root elongation (Lactuca sativa). EPA 600/3-88/029. Washington, DC: U.S. Environmental Protection AgencyGoogle Scholar
  15. EPA (2017). PBTprofiler Accessed 6 February 2017.
  16. Federle, T. W., Kaiser, S. K., & Nuck, B. A. (2002). Fate and effects of triclosan in activated sludge. Environmental Toxicology Chemistry, 21(7), 1330–1337. Scholar
  17. Fortunato, M. S., Fuentes Abril, N. P., Martinefsky, M., Trípodi, V., Papalia, M., Rádice, M., et al. (2016). Aerobic degradation of ibuprofen in batch and continuous reactors by an indigenous bacterial community. Environmental Technology, 37(20), 2617–2626. Scholar
  18. Gaze, W. H., Abdouslam, N., Hawkey, M., & Wellington, E. M. H. (2005). Incidence of class 1 integrons in a quaternary ammonium compound-polluted environment. Antimicrobial Agents and Chemotherapy, 49(5), 1802–1807. Scholar
  19. González, O., Carme Sans, C., & Esplugas, S. (2007). Sulfamethoxazole abatement by photo-Fenton: toxicity, inhibition and biodegradability assessment of intermediates. Journal of Hazardous Materials, 146(3), 459–464. Scholar
  20. Hall-Stoodley, H., Costerton, J. W., & Stoodley, P. (2004). Bacterial biofilms: from the natural environment to infectious diseases. Nature Reviews Microbiology, 2(2), 97–108. Scholar
  21. Hernando, M. D., Fernández Alba, A. R., Tauler, R., & Barceló, D. (2005). Toxicity assays applied to wastewater treatment. Talanta, 65(2), 358–366. Scholar
  22. ISO 11348-3 (1998). Water quality. Determination of the inhibitory effect of water samples on the light emission of Vibrio fischeri (Luminescent bacteria test). Geneva: SwitzerlandGoogle Scholar
  23. ISO 8692 (2004). Water quality. Freshwater algal growth inhibition test with unicellular green algae. Polski Komitet Normalizacyjny: PolskaGoogle Scholar
  24. Ivanković, T., & Hrenović, J. (2010). Surfactants in the environment. Archives of Industrial Hygiene and Toxicology, 61(1), 95–110. Scholar
  25. Jesus, F. T., Oliveira, R., Silva, A., Catarino, A. L., Soares, A. M. V. M., Nogueira, A. J. A., et al. (2013). Lethal and sub lethal effects of the biocide chlorhexidine on aquatic organisms. Ecotoxicology, 22(9), 1348–1358. Scholar
  26. Jones, O. A. H., Voulvoulis, N., & Lester, J. N. (2001). Human pharmaceuticals in the aquatic environment a review. Environmental Technology, 22(12), 1383–1394. Scholar
  27. Khan, A. H., Topp, E., Scott, A., Sumarah, M., Macfie, S. M., & Ray, M. B. (2015). Biodegradation of benzalkonium chlorides singly and in mixtures by a Pseudomonas sp. isolated from returned activated sludge. Journal of Hazardous Materials, 299, 595–602. Scholar
  28. Koäljalg, S., Naaber, P., & Mikelsaar, M. (2002). Antibiotic resistance as an indicator of bacterial chlorhexidine susceptibility. Journal of Hospital Infection, 51(2), 106–113. Scholar
  29. Korol, S., Orsingher, M., Santini, P., Moretton, J., & D’Aquino, M. (1989). Biodegradation of phenolic compounds, II. Effects of inoculum, xenobiotic concentration and adaptation on Acinetobacter and Pseudomonas phenol degradation. Revista Latinoamericana de Microbiología, 31(2), 117–120.Google Scholar
  30. Kümmerer, K., Eitel, A., Braun, U., Hubner, P., Daschner, F., Mascart, F., et al. (1997). Analysis of benzalkonium chloride in the effluent from European hospitals by solid-phase extraction and high-performance liquid chromatography with post-column ion-pairing and fluorescence detection. Journal of Chromatography A, 774(1–2), 281–286. Scholar
  31. Kümmerer, K. (2001a). Drugs in the environment: emission of drugs, diagnostic aids and disinfectants into wastewater by hospitals in relation to other sources. Chemosphere, 45(3), 957–969. Scholar
  32. Kümmerer, K. (2001b). Emission and biodegradability of pharmaceuticals, contrast media, disinfectants and AOX from hospitals. In K. Kümmerer (Ed.), Pharmaceuticals in the Environment- Sources, fate, effects and risk (pp. 29–41). Heidelberg: Springer-Verlag.CrossRefGoogle Scholar
  33. Kümmerer, K., Alexy, R., Hüttig, J., & Schöll, A. (2004). Standardized tests fail to assess the effects of antibiotics on environmental bacteria. Water Research, 38(8), 2111–2116. Scholar
  34. Kümmerer, K. (2004). Resistance in the environment. Journal of Antimicrobial Chemotherapy, 54(2), 311–320. Scholar
  35. Kümpel, T., Alexy, R., & Kümmerer, K. (2001). What we know about antibiotics in the environment? In K. Kümmerer (Ed.), Pharmaceuticals in the Environment- Sources, fate, effects and risk (pp. 67–76). Heidelberg: Springer-Verlag.CrossRefGoogle Scholar
  36. Labranche, L. P., Dumont, S. N., Levesque, S., & Carrier, A. (2007). Rapid determination of total benzalkonium chloride content in ophthalmic formulation. Journal of Pharmaceutical and Biomedical Analysis, 43(3), 989–993. Scholar
  37. La Farré, M., Pérez, S., Kantiani, L., & Barcelo, D. (2008). Fate and toxicity of emerging pollutants, their metabolites and transformation products in the aquatic environment. Trends in Analytical Chemistry, 27(11), 992–1007. Scholar
  38. Lara Martín, P. A., Li, X., Bopp, R. F., & Brownawell, B. J. (2010). Occurrence of alkyltrimethylammonium compounds in urban estuarine sediments: behentrimonium as a new emerging contaminant. Environmental Science & Technology, 44(19), 7569–7575. Scholar
  39. Li, X., Luo, X., Mai, B., Liu, J., Chen, L., & Lin, S. (2014). Occurrence of quaternary ammonium compounds (QACs) and their application as a tracer for sewage derived pollution in urban estuarine sediments. Environmental Pollution, 185, 127–133. Scholar
  40. Loveira, E., Fiol, P., Senn, A., Curutchet, G., Candal, R., & Litter, M. (2012). TiO2-photocatalytic treatment coupled with biological systems for the elimination of benzalkonium chloride in water. Separation and Purification Technology, 91, 108–116. Scholar
  41. Lowry, J. B. (1979). Direct spectrophotometric assay of quaternary ammonium compounds using bromothymol blue. Journal of Pharmaceutical Sciences, 68(1), 110–111. Scholar
  42. Martínez-Carballo, E., Sitka, A. L., Gonzalez-Barreiro, C., Kreuzinger, B. N., Fürhackerc, M., Scharf, S., et al. (2007a). Determination of selected quaternary ammonium compounds by liquid chromatography with mass spectrometry. Part I. Application to surface, waste and indirect discharge water samples in Austria. Environmental Pollution, 145(2), 489–496. Scholar
  43. Martínez-Carballo, E., González Barreiro, C., Sitka, A., Kreuzinger, N., Scharf, S., & Gans, O. (2007b). Determination of selected quaternary ammonium compounds by liquid chromatography with mass spectrometry. Part II. Application to sediment and sludge samples in Austria. Environmental Pollution, 146(2), 543–547. Scholar
  44. Morrall, D., McAvoy, D., Schatowitz, B., Inauen, J., Jacob, M., Hauk, A., et al. (2004). A field study of triclosan loss rates in river water (Cibolo Creek, TX). Chemosphere, 54(5), 653–660. Scholar
  45. Natale, O. E. (2005). Water quality indicators for the La Plata River basin. Vienna: IWG-Environmental International Work Session on Water Statistics Accessed 6 March 2017.Google Scholar
  46. Neumegen, R. A., Fernández-Alba, A. R., & Chisti, Y. (2005). Toxicities of triclosan, phenol, and copper sulfate in activated sludge. Environmental Toxicology, 20(2), 160–164. Scholar
  47. Nishihara, T., Okamoto, T., & Nishiyama, N. (2000). Biodegradation of didecyldimethylammonium chloride by Pseudomonas fluorescens TN4 isolated from activated sludge. Journal of Applied Microbiology, 88(4), 641–647. Scholar
  48. OECD 301D. (1992). Guidelines for testing of chemicals. Closed bottle test. Paris: Organisation of Economic Cooperation and Development.Google Scholar
  49. Patrauchan, M. A., & Oriel, P. J. (2003). Degradation of benzyldimethylalkylammonium chloride by Aeromonas hydrophila sp K. Journal of Applied Microbiology, 94(2), 266–272. Scholar
  50. Prince, S. J., McLaury, H. J., Allen, L. V., & McLaury, P. (1999). Analysis of benzalkonium chloride and its homologs: HPLC versus HPCE. Journal of Pharmaceutical and Biomedical Analysis, 19(6), 877–882. Scholar
  51. Rosal, R., Rodríguez, A., Perdigón-Melón, J. A., Petre, A., García-Calvo, E., Gómez, M. J., et al. (2010). Occurrence of emerging pollutants in urban wastewater and their removal through biological treatment followed by ozonation. Water Research, 44(2), 578–588. Scholar
  52. Salibián, A. (2006). Ecotoxicological assessment of the highly polluted Reconquista River of Argentina. In G. W. Ware, H. N. Nigg, & D. R. Dorge (Eds.), Reviews of environmental contamination and toxicology (pp. 35–65). New York: Springer.CrossRefGoogle Scholar
  53. Sütterlin, R., Coker, A., & Kümmerer, K. (2008a). Mixtures of quaternary ammonium compounds and anionic organic compounds in the aquatic environment: elimination and biodegradability in the closed bottle test monitored by LC–MS/MSH. Chemosphere, 72(3), 479–484. Scholar
  54. Sütterlin, H., Alexy, R., & Kümmerer, K. (2008b). The toxicity of the quaternary ammonium compound benzalkonium chloride alone and in mixtures with other anionic compounds to bacteria in test systems with Vibrio fischeri and Pseudomonas putida. Ecotoxicology and Environmental Safety, 71(2), 498–505. Scholar
  55. Tandukar, M., Oh, S., Tezel, U., Konstantinidis, K. T., & Pavlostathis, S. G. (2013). Long-term exposure to benzalkonium chloride disinfectants results in change of microbial community structure and increased antimicrobial resistance. Environmental Science Technology, 47(17), 9730–9738. Scholar
  56. Tezel, U., & Pavlostathis, S. G. (2009). Transformation of benzalkonium chloride under nitrate reducing conditions. Environmental Science Technology, 43(5), 1342–1348. Scholar
  57. Xia, K., Bhandari, A., Das, K., & Pillar, G. (2005). Occurrence and fate of pharmaceuticals and personal care products (PPCPs) in biosolids. Journal of Environmental Quality, 34(1), 91–104. Scholar
  58. Ying, G. G. (2006). Fate, behavior and effects of surfactants and their degradation products in the environment. Environmental International, 32(3), 417–431. Scholar
  59. Zhang, C., Tezel, U., Li, K., Liu, D., Ren, R., Du, J., et al. (2011). Evaluation and modeling of benzalkonium chloride inhibition and biodegradation in activated sludge. Water Research, 45(3), 1238–1246. Scholar
  60. Zhang, C., Cui, F., Zeng, G. M., Jiang, M., Yang, Z. Z., Yu, Z. G., et al. (2015). Quaternary ammonium compounds (QACs): a review on occurrence, fate and toxicity in the environment. Science of the Total Environment, 518-519, 352–362. Scholar

Copyright information

© Springer International Publishing AG, part of Springer Nature 2018

Authors and Affiliations

  • María Susana Fortunato
    • 1
  • Sabina Baroni
    • 1
  • Ana Julieta González
    • 1
    • 2
  • Jesús David Álvarez Roncancio
    • 1
  • Mariana Papalia
    • 3
  • Manuela Martinefsky
    • 4
  • Valeria Trípodi
    • 2
    • 3
    • 4
  • Estela Planes
    • 5
  • Alfredo Gallego
    • 1
  • Sonia Edith Korol
    • 1
  1. 1.Facultad de Farmacia y Bioquímica, Cátedra de Salud Pública e Higiene AmbientalUniversidad de Buenos AiresBuenos AiresArgentina
  2. 2.Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET)Buenos AiresArgentina
  3. 3.Facultad de Farmacia y Bioquímica, Laboratorio de Resistencia Bacteriana, Cátedra de MicrobiologíaUniversidad de Buenos AiresBuenos AiresArgentina
  4. 4.Facultad de Farmacia y Bioquímica, Departamento de Tecnología, FarmacéuticaUniversidad de Buenos AiresBuenos AiresArgentina
  5. 5.Centro de Química y Petroquímica (CEQUIPE)Instituto Nacional de Tecnología IndustrialSan MartínArgentina

Personalised recommendations